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Bam35, a 15-kbp double-stranded DNA phage, infects Bacillus thuringiensis. Recently, sequencing of the related Bacillus cereus revealed a 15.1-kbp linear plasmid, pBClin15. We show that pBClin15 closely resembles Bam35 and demonstrate conversion of Bam35 to a prophage. This state is common, as several B. thuringiensis strains release Bam35-related viruses.Phage Bam35 was originally isolated from Bacillus thuringiensis var. alesti strain 35 (1). As Bam35 could also be isolated from Bam35-resistant colonies of Bacillus megaterium and from a sensitive strain of B. thuringiensis var. entomocidus, a carrier state has been proposed (1). Recently, a clear-plaque mutant (Bam35c) was sequenced and further characterized (25). This work showed that Bam35 is related to PRD1, a linear double-stranded DNA virus with 5Ј-terminal proteins and an internal lipid membrane (4). PRD1 infects gram-negative bacteria harboring an IncP, -W, or -N conjugative plasmid. Structural analyses of PRD1 have, surprisingly, indicated that this member of the Tectiviridae is similar to the Adenoviridae, Phycodnaviridae, and Iridoviridae, which all infect eukaryotic hosts (9,24,33). This has led to the hypothesis that all these viruses belong to the same lineage, with a common ancestor existing before the separation of the three domains of life (5, 6).Phage Bam35 is closely related to Bacillus cereus plasmid pBClin15. Bacilli harbor a large variety of plasmids, which include a linear species of Ϸ15 kb (2,13,15,31,34). Recently, the genome sequencing of Bacillus cereus ATCC 14579 revealed the sequence of a linear plasmid (pBClin15) of 15,100 bp (19). While studying Bam35, we matched putative genes of the pBClin15 plasmid in independent database searches for the Bam35c coat protein and ATPase. After submission of this note, the nucleotide sequence of a linear B. thuringiensis plasmid, pGIL01, which differs from the Bam35c sequence by only approximately 10 nucleotides was published (32).To investigate the relationship of pBClin15 to Bam35c (14,935 bp), their genomes were compared. The organizations of the open reading frames (ORFs) of pBClin15 and those of Bam35c are highly similar (Fig. 1). Moreover, their sequences agree with 45 to 81% identity. The corresponding amino acids have 18 to 88% identity ( Table 1). The gene identification of Bam35c (25) was based on the reasonably detailed understanding of the corresponding PRD1 genes (7,8,16). A similar annotation shows that pBClin15 is related to Bam35 and PRD1 (Table 1). Among the most conserved genes in pBClin15 are those assigned to viral capsid components. These include the major coat protein 15 (corresponding to PRD1 protein P3) and the unique vertex packaging proteins 12, 14, and 16 (proteins P9, P20, and P22, respectively, in PRD1). Interestingly, the most conserved protein corresponds to a LexA-type transcription regulator (17, 21) homologue found in Bam35 but absent in PRD1.We then explored the relationship between pBClin15, Bam35c, and PRD1 by comparing their major coat proteins. First, the th...

Icosahedral double-stranded DNA (dsDNA) bacterial viruses are known to package their genomes into preformed procapsids via a unique portal vertex. Bacteriophage PRD1 differs from the more commonly known icosahedral dsDNA phages in that it contains an internal lipid membrane. The packaging of PRD1 is known to proceed via preformed empty capsids. Now, a unique vertex has been shown to exist in PRD1. We show in this study that this unique vertex extends to the virus internal membrane via two integral membrane proteins, P20 and P22. These small membrane proteins are necessary for the binding of the putative packaging ATPase P9, via another capsid protein, P6, to the virus particle.The genome packaging of icosahedral double-stranded DNA (dsDNA) bacteriophages occurs by translocation of the genome into preformed procapsids. This translocation is performed by a specific enzyme, a terminase or a packaging ATPase, and is powered by ATP hydrolysis. The packaging of DNA occurs at a unique portal vertex, which also functions as the head-to-tail connector as well as the channel through which DNA is injected into the host cell during infection. The portals characterized thus far are ring-like structures of portal protein multimers with a central channel correctly sized for the dsDNA to be threaded through. Packaging is believed to occur by winding the DNA up through this central channel, involving movement of the portal structure (16,20,35,42,43,64,66). A symmetry mismatch between the packaging machinery and the rest of the capsid has been proposed to assist the movement of the packaging machinery with respect to the capsid during nucleic acid transport (35,42,64).The most detailed description of a packaging system exists for bacterial virus 29, a short-tailed icosahedral prolate virus with a linear dsDNA genome, infecting the gram-positive Bacillus subtilis (2). The 29 portal occupying a pentagonal vertex is a dodecamer of portal protein gp10, forming a propellershaped structure with a central channel (35,64). In addition to the portal, which is attached to the prohead, the packaging machinery consists of five or six copies of the viral ATPase (gp16) and six copies of a 29-encoded packaging RNA (pRNA) (36-39, 74). The DNA, connector, and proheadpRNA-ATPase complex form a set of concentric structures with 10-, 12-, and 5-or 6-fold symmetry, respectively, embedded in the 5-fold vertex of the prohead (35,64).A portal assembly complex has also been identified in a eukaryotic virus. The portal of herpes simplex virus type 1 (HSV-1), identified by immunogold labeling, is located at a single vertex of the icosahedral HSV-1 capsid and contains a ring-shaped multimer of the UL6 protein (52). So far, other portal vertices of icosahedral eukaryotic viruses have not been described. It is conceivable, however, that many more complex icosahedral viruses assemble through packaging of empty precursor particles, which would require a unique portal complex.PRD1 is the type organism for the family Tectiviridae (7). It is a bacterial virus that...